Down syndrome is a significant social and medical problem. As a cause of severe mental retardation, it affects greater than 0.1% of live births, and in addition, is associated with numerous developmental defects, a very high incidence of leukemia, and the eventual development of the neuropathology of Alzheimer's disease. Understanding the pathogenesis of Down syndrome may allow amelioration of its effects, and should assist in the understanding of the associated problems. As it is a genetic disease, a detailed description is necessary of the region of the human genome responsible for the Down syndrome phenotype. This proposal involves a new approach to the isolation and characterization of this material. Down syndrome is known to result from trisomy of the long arm of chromosome 21, a segment of the human genome comprising approximately 30 million base pairs of DNA. There is much evidence that genes in this region are actively transcribed, and many indications that products of these genes are directly relevant to many Down syndrome associated abnormalities. However, it is also becoming increasingly clear that many human genes are too large (>40-50kb) to be cloned as functional fragments in single lambda or cosmid clones. A detailed understanding of the Down syndrome region may require information regarding interactions between genes, or their coordinate regulation. To do this, it is necessary to develop techniques for the cloning and manipulation of fragments of DNA spanning 200 to possibly 1000kb or more. The specific aims of this proposal involve: i) construction of chromosome 21 DNA libraries of large insert size (>200kb) using an S. cerevisiae-host/vector system. ii) analysis of the library inserts by pulsed field gel electrophoresis to determine their sequence content and organization; and iii) transfer of the cloned DNA in these libraries to mammalian cells by their fusion with protoplasts of the yeast host. It is intended that these techniques will help to determine the number and identity of genes and their organization, in several interesting regions of chromosome 21. This information will complement that available from conventional library methods.